PROGRESS IN FP5: Pre-Breeding and Trait Discovery – 2021

Drought screening was done in the screenhouse at IITA for cowpea crop wild relatives (CWR) at seedling stage using wooden box technique (Singh et al. 1999). The same set of genotypes was screened for terminal drought in Minjibir (Nigeria). Several cowpea genotypes showed tolerance to drought at seedling stage with more than 75% recovery rate, as well as those with terminal drought tolerance. From these, three genotypes (TVNu-2158, TVNu-2159 and TVNu2192) showed consistent results for seedling stage and terminal drought screening. The consistent cowpea CWRs were crossed with some of the elite cultivated varieties from INERA-Burkina Faso, INRAN-Niger, and IITA-Nigeria and incorporated into pre-breeding resources. About 40 F5 lines derived from these crosses will be characterized for terminal drought tolerance.

IITA also received 63 pre-bred soybean lines from the Colombian private sector, Semillas Panorama and introduced them to the Nigerian and Zambian soybean breeding programs.  In Nigeria, four pre-bred Colombian lines yielded over 2 ton/ha, exceeding the performance of the most popular variety. In Zambia, four of the pre-bred soybean lines performed well both under normal and reduced phosphorus application. Based on these trials, most of the pre-bred soybean lines were found to be better adapted to Nigeria than to Southern Africa with a potential to contribute enormously to the West African soybean breeding community. While 205 soybean accessions were received from USA through the Soybean Innovation Lab, their full evaluation will be done in Lusaka and the highlands environment in Kenya.

At CERAAS in Senegal, two AB-QTLs of groundnut populations from the cross between Fleur11 and (A. ipaensis x A. correntina) 4x, comprising of 104 BC1F4 and 115 BC2F3 were characterized at the phenotypic and genotypic levels. About 219 individuals from the AB-QTL population were phenotyped along with Fleur11 and two checks during the rainy-season of 2021 in Nioro (Senegal) for several traits including disease resistance early leaf spot (ELS) and late leaf spot (LLS) diseases and yield component traits (pod and haulm weight, pod and seed size, maturity etc.). Leveraging on initiatives of the Peanut Innovation Lab in USA, this population was also genotyped using the Axiom_Arachis V2 SNP chip. Following the ongoing analysis of phenotypic and genotypic data, QTL mapping and trait marker discovery will be carried out.

In pigeonpea, QTL-seq approach was used to identify candidate genes by genotyping pools of fertile and sterile F2s derived from ICPA 2039 × ICPL 87119. To develop NILs and high-resolution mapping of fertility restoration in pigeonpea, BC1F2s (derived from ICPA 2039 x ICPL 87119) were selected and crossed with male sterile parent (ICPA 2039) to generate F1s. A total of 122 F1s were developed and planted in the field that were also tested with fertility restoration associated RfQ1 and RfQ4 markers to select true heterozygote to be selfed with the harvested F2 seeds.

To develop new genetic stocks and enable high resolution mapping of target traits in pigeonpea, MAGIC and NAM populations were used to phenotype in the fields in as well as at different NARS centers. These populations are also being genotyped with EGS markers for creating a sub-set with favourable alleles for must have traits including resistance to fusarium wilt (FW) and sterility mosaic disease (SMD). Besides, to develop FW and SMD resistant lines in pigeonpea, a total of 341 lines of 12 crosses (8 in BC2F1 and 4 in BC1F1) were subjected to foreground selection with 20 markers (10 for FW and 10 for SMD). Lines carrying favourable alleles for FW and SMD resistance were selected where the selected BC2F1s will be used for second round of backcrossing, and selfed to develop BC2F2s for the background and foreground selections.

In chickpea, extreme bulks for 100 seed weight, days to 50% flowering and days to maturity were prepared and sequenced along with parental lines at 10X coverage. Identification of SNPs affecting the trait is in progress. Four parental chickpea genotypes that are parents of two near isogenic lines (NIL) contrasting for FW resistance were whole genome sequenced leading to the identification of candidate genes explaining complex mechanisms associated the resistance. Based on collaborative work on improving chickpea mega varieties, two lines developed by introgressing QTL-hotspot in the background of JG11 and RSG888 were promoted to AVT 2 and four introgression lines in the background of JAKI 9218 and DCP 92-3 have been nominated for AVT 1 trials of AICRP-Chickpea for possible release for commercial cultivation in India.  To understand differentially expressed and methylated regions under drought stress in chickpea genome, four samples representing two drought tolerant genotypes (ICC 4958 and ICC 8261), and two drought susceptible genotypes (ICC 1882 and ICC 283) were targeted that resulted in the identification of 592 unique chickpea specific miRNA and 12 novel miRNAs which will be further characterized for their role in drought/stress tolerance. For salinity tolerance, population of a recombinant inbred line (RIL) developed using parental lines ICCV 10 (salt tolerant) and DCP 92-3 (salt-sensitive) was genotyped using Axiom®CicerSNP array to construct a linkage map comprising 1856 SNP markers spanning 1106.3 cM. Extensive analysis of the phenotyping and genotyping data identified 28 QTLs explaining up to 28.40% of the phenotypic variance in the population. QTL clusters on CaLG03 and CaLG06, each harboring major QTLs for yield and yield component traits under salinity stress Trait linked markers are being validated for deploying in the breeding program were identified. In addition, 280 accessions from chickpea reference set were evaluated for 11 nutrition traits and used for GWAS, resulting in the detection of 119 Marker Trait associations (MTAs) for 11 out of 12 targeted traits that are currently being validated. For AB resistance in chickpea, transcriptome, small RNA and degradome sequencing resulted in the identification of 6767 differentially expressed genes, 297 miRNAs related to pathogenesis‐related proteins and disease resistance genes. The combined analysis of both small RNA and transcriptome data identified 12 miRNA‐mRNA interaction pairs related to AB infection.

In cowpea, validation of SNP markers identified from GWAS for Striga resistance is underway where RILs have been developed from two Striga resistant cowpea parents that are currently being used to validate the Striga resistant QTLs identified through GWAS. Candidate SNPs are being converted to KASP markers, followed by validation in RIL populations for eventual deployment in marker-assisted selection.

To map genomic regions for flour keeping quality in pearl millet, 81 pearl millet inbred germplasm association panel (PMiGAP) lines have been evaluated for acid value (AV) and sensory parameters (taste and aroma) at three different time intervals. More data is being generated on the diversity panel from grain samples in replicated trials from multi environments. These data will be used to carry out a genome-wide scan for favorable alleles leading to enhanced flour keeping quality.  Four SNP panel marker for grain iron and zinc content in pearl millet were also developed, verified and validated. The SNPs were validated in diverse lines from the Indian and African breeding programs that are being effectively used to reject low Fe and Zn lines. For further characterization of root traits, pearl millet backcross inbred lines (BC2F4) for two root traits (early primary root growth and soil aggregation – 180 lines/trait) were genotyped by sequencing. Phenotyping for soil aggregation of the panel of lines are underway. New field trials confirmed the correlation between early primary root growth and tolerance to drought stress post germination in pearl millet (crop establishment).

In sorghum, 46 genotypes were evaluated for drought tolerance and striga resistance that were characterized using Diversity Arrays Technology (DArT) sequencing. A new source of drought tolerance (Lodoka) and Striga resistance (ICSV III IN and F6YQ212) was also identified for further trait characterization and mapping of the responsible QTLs.

Enabling technologies mainly focused on establishing protocols for proof-of-concept in genome editing, second-generation transformation (QuickCrop from Corteva), systematic mutant population, phenotypic screening protocols, and rapid generation turnover (RGT) with the following major achievements:

  • Collaboration with CIMMYT to develop specialized nethouse facility for phenotyping resistance to Fall armyworm in sorghum, millet and maize was completed at ICRISAT, Patancheru, Hyderabad.
  • All field experiments were designed using BMS in 2020. In cowpea, all main field experiments which included Initial Evaluation Trial (IET), Preliminary Variety Trial (PVT), Advanced Variety Trial (AVT), Cowpea International Trial (CIT) and Regional Coordinated Trials were designed using BMS. For these trials, electronic field books were generated and are ready for uploading into tablets to capture observation data during the current cropping season.
  • Data for all ICRISAT mandate crops were loaded on GOBii and complete DNA sampling workflow was done by connecting BMS and GOBii.
  • In collaboration with Corteva Agriscience, QuickCrop expression constructs for gene editing were developed for efficient regeneration and transformation for gene editing. Gene editing is being carried out in elite backgrounds of sorghum for accelerated trait deployment.
  • Using reverse genetics approaches, a TILLING population of sorghum was characterized for strigolactone exudates and validated for primary strigolactone pathway genes in two lines. The selected mutant lines were advanced to T1 generation and sent to Niger and Mali to evaluate their efficacy under Striga infestation.
  • Transformative RapidGen methodologies for accelerated breeding cycles in cowpea have been developed using 10 genotypes. Overall, 4-6 generations may be taken in a year, thereby offering tremendous opportunities to enhance the rate of genetic gain and accelerate breeding pipelines for this important crop.
  • To strengthen the capacity in climate change and plant protection research, collaboration with advance research institutions such as University of Strathclyde, Scotland, and Wageningen University Research Netherlands has been established. This will enable the development of high throughput phenotyping platforms using sensor-based technology for rapid detection of pathogens and develop modelling tools to predict the potential distribution of pests and diseases under future climate scenarios.
  • A workshop for early career researchers was conducted during the annual general assembly of TIGR2ESS from 20 to 24 January at ICRISAT, HQ (50 participants). The 15th training course and International Workshop on “Next Generation Genomics for Developing Climate Resilient Agriculture” was also conducted during 10-15 February at the ICRISAT headquarters (60 participants). Other training programs included a Virtual webinar in the series of Next Generation Genomics and Integrated Breeding for Crop Improvement on Genomics for food, health and nutrition, 14 May 2020 (~3000 virtual participants).
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